Discovery of a first-in-class protein degrader for the c-ros oncogene 1 (ROS1).

The c-ros oncogene 1 (ROS1), an oncogenic driver, is known to induce non-small cell lung cancer (NSCLC) when overactivated, particularly through the formation of fusion proteins. Traditional targeted therapies focus on inhibiting ROS1 activity with ROS 1 inhibitors to manage cancer progression. However, a new strategy involving the design of protein degraders offers a more potent approach by completely degrading ROS1 fusion oncoproteins, thereby effectively blocking their kinase activity and enhancing anti-tumour potential. Utilizing PROteolysis-TArgeting Chimera (PROTAC) technology and informed by molecular docking and rational design, we report the first ROS1-specific PROTAC, SIAIS039. This degrader effectively targets multiple ROS1 fusion oncoproteins (CD74-ROS1, SDC4-ROS1 and SLC34A2-ROS1) in engineered Ba/F3 cells and HCC78 cells, demonstrating anti-tumour effects against ROS1 fusion-driven cancer cells. It suppresses cell proliferation, induces cell cycle arrest, and apoptosis, and inhibits clonogenicity. The anti-tumour efficacy of SIAIS039 surpasses two approved drugs, crizotinib and entrectinib, and matches that of the top inhibitors, including lorlatinib and taletrectinib. Mechanistic studies confirm that the degradation induced by 039 requires the participation of ROS1 ligands and E3 ubiquitin ligases, and involves the proteasome and ubiquitination. In addition, 039 exhibited excellent oral bioavailability in a mouse xenograft model, highlighting its potential for clinical application. In conclusion, our study presents a promising and novel therapeutic strategy for ROS1 fusion-positive NSCLC by targeting ROS1 fusion oncoproteins for degradation, laying the foundation for the development of further PROTAC and offering hope for patients with ROS1 fusion-positive NSCLC.

Utilization of macrocyclic peptides to target protein-protein interactions in cancer.

Protein-protein interactions (PPIs) play vital roles in normal cellular processes. Dysregulated PPIs are involved in the process of various diseases, including cancer. Thus, these PPIs may serve as potential therapeutic targets in cancer treatment. However, despite rapid advances in small-molecule drugs and biologics, it is still hard to target PPIs, especially for those intracellular PPIs. Macrocyclic peptides have gained growing attention for their therapeutic properties in targeting dysregulated PPIs. Macrocyclic peptides have some unique features, such as moderate sizes, high selectivity, and high binding affinities, which make them good drug candidates. In addition, some oncology macrocyclic peptide drugs have been approved by the US Food and Drug Administration (FDA) for clinical use. Here, we reviewed the recent development of macrocyclic peptides in cancer treatment. The opportunities and challenges were also discussed to inspire new perspectives.

Research on antibody changes and nucleic acid clearance in COVID-19 patients treated with convalescent plasma.

PURPOSE: To investigate changes in the production of IgM and IgG antibodies and the negative transformation of viral nucleic acids in COVID-19 patients after convalescent plasma therapy, and also to discuss the clinical therapeutic effect, so as to provide a basis for the treatment of COVID-19 using specific antibodies. METHODS: The convalescent plasma of recovered patients from COVID-19 was used to treat other patients, and the levels of antibodies IgM and IgG and the nucleic acid genes ORF1ab and N in the patients were tested regularly for statistical comparison and analysis. RESULTS: In general, the Ct value and concentration of IgM and IgG antibodies in the plasma infusion group were significantly higher (1-3 times higher) than those in the non-plasma infusion group, respectively, but these differences were not significant (P>0.05). However, the content of antibodies in severe patients in the plasma transfusion group was significantly higher than those in the non-plasma transfusion group at discharge, the results being statistically significant (P<0.05). CONCLUSIONS: The application of convalescent plasma significantly increases the antibody content in severe and critical inpatients, effectively enhances immune function, accelerates the clearance of virus and the nucleic acid negative conversion rate, and significantly promotes early improvement in COVID-19 patients.

Two novel strategies to overcome the resistance to ALK tyrosine kinase inhibitor drugs: Macrocyclic inhibitors and proteolysis-targeting chimeras.

Lung cancer is the most malignant tumor in the worldwide. About 3%-5% non-small cell lung cancer (NSCLC) patients carry anaplastic lymphoma kinase (ALK) gene fusions and receive great benefits from ALK-targeted therapy. However, drug resistance inevitably occurs even with the most potent inhibitor drug lorlatinib. About half of the resistance are caused by alteration in ALK proteins for earlier ALK TKI drugs and near one-third of loratinib resistant cases are caused by compound mutations without current effective treatment strategy in clinic. Novel strategies are in great need to overcome drug resistance. Lately, two novel strategies have been developed and attracted great attentions for their potentials to overcome drug resistance problems: (1) developed small compact macrocyclic ALK kinase inhibitors and (2) developed ALK targeted proteolysis-targeting chimera (PROTAC) drugs. The macrocyclic molecules are small and compact in size, brain barrier permeable, and highly potent against lorlatinib-resistant compound mutations. Developed ALK targeted PROTAC molecules could degrade oncogenic ALK driver proteins. Some showed superiority in killing ALK positive cancer cells and inhibiting the growth of cells expressing G1202R resistant ALK proteins comparing to inhibitor drugs. The update on these two treatment strategies was reviewed.

Effective degradation of EGFRL858R+T790M mutant proteins by CRBN-based PROTACs through both proteosome and autophagy/lysosome degradation systems.

Targeted therapy of treating patients with specific tyrosine kinase inhibitors (TKIs) is currently the standard care for epidermal growth factor receptor (EGFR) mutant non-small cell lung cancer. However, the inevitably developed drug resistance in patients to EGFR TKIs is the biggest obstacle for cancer targeted therapy. About 60% of drug resistance to the 1st generation of EGFR TKIs was resulted from an acquired T790M mutation in the kinase domain of EGFR protein. Proteolysis targeting chimera (PROTAC) is a lately-developed technology to target point of interest proteins for degradation. Because EGFR-mutant lung cancers are highly dependent on EGFR proteins, designing specific PROTAC molecules to degrade EGFR proteins from cancer cells provides a very promising strategy to treat such patients and eradicate drug resistance. Currently, there is no cereblon (CRBN)-based PROTAC reported able to degrade T790M-containing EGFR resistant proteins. In this study, we synthesized two novel CRBN-based EGFR PROTACs, SIAIS125 and SIAIS126, based on EGFR inhibitor canertinib and cereblon ligand pomalidomide. These two degraders displayed potent and selective antitumor activities in EGFR TKI resistant lung cancer cells. Firstly, they could selectively degrade EGFRL858R+T790M resistant proteins in H1975 cells at the concentration of 30-50 nM, and EGFREx19del proteins in PC9 cells. But they did not degrade EGFREx19del+T790M mutant proteins in PC9Brca1 cells or wild type EGFR in A549 lung cancer cells. They could also selectively inhibit the growth of EGFR mutant lung cancer cells but not that of normal cells or A549 cells. Secondly, the degradation of EGFRL858R+T790M proteins was long lasting up to 72 h. Thirdly, these degraders displayed better inhibition of EGFR phosphorylation in H1975 cells and PC9Brca1 cells comparing to canertinib. Finally, these degraders could also induce significant apoptosis and cell cycles arrest in H1975 cells. Pre-incubation with canertinib, pomalidomide or ubiquitination inhibitor MLN4924 totally blocked EGFR degradation by PROTACs. Mechanistic studies showed that PROTAC could induce autophagy in lung cancer cells. PROTAC-induced EGFR degradation acted through both ubiquitin/proteosome system and ubiquitin/autophagy/lysosome system. Elevating autophagy activities enhanced EGFR degradation and cell apoptosis induced by PROTACs. Our research not only offered a novel PROTAC tool to target EGFR TKI drug resistance in lung cancer, but also firstly demonstrated that the involvement of autophagy/lysosome system in PROTAC- mediated target protein degradation.

Structure-based discovery of SIAIS001 as an oral bioavailability ALK degrader constructed from Alectinib.

Fusion proteins of the anaplastic lymphoma kinase (ALK) are promising therapeutic targets for cancer and other human diseases, especially for non-small cell lung cancer (NSCLC) and anaplastic large-cell lymphomas (ALCLs). We described herein a structure-based design, synthesis, and evaluation of ALK PROTACs (proteolysis-targeting chimeras) based on Alectinib as the warhead. We firstly screened CRBN ligands as the E3 ligase moiety, then obtained a series of potent ALK degraders based on different CRBN ligands, exemplified by SIAIS091 and SIAIS001 with lenalidomide/thalidomide-based linkers. Both of them induced effective ALK degradation at low nanomolar concentrations in cells, and showed much better growth inhibition effects than Alectinib. SIAIS091 or SIAIS001 also promoted cell cycle arrest in G1/S phase. Finally, SIAIS001 exhibited good oral bioavailability in Pharmacokinetics study.

Enhancing capacity building to climate adaptation and water conservation among Chinese young people.

Capacity development has been regarded as one of many measures to empower the abilities of nations to adapt to a changing climate. Promoting public engagement in water conservation, especially among young people since they will be leading decision making in the near future, is one effective strategy for adapting to the changing climate. This research presented a water footprint adaptation (WF) approach which attempted to link climate adaptation and capacity development with saving water strategy. The approach was tested in cooperation with two universities in Shaanxi province aiming to develop a starting point for WF evaluation and develop an improvement response. The results showed that the college students in our samples reduced their direct and indirect water footprints by 27.39% and 6.50%, respectively, in the post-intervention phase. The evaluation of the improvements proved that the WF approach to be efficient, the awareness of the college students on the matter could be increased. Additionally, the findings of the research indicated that the college students became change agent, expressing the desire to act as multipliers and to help the movement and spread of important knowledge about methods for alleviating water stress and about vulnerability to the changing climate. As expected, the awareness of water scarcity and perceptions of climate change had statistically significant effects on the water footprints, which was consistent with our hypothesis. Our approach helped participants develop capacity by revealing the linkage between their local level actions and the various aspects of adaptation to changing climate at the global level. This strategy will provide a comparative basis for water policy makers to adopt appropriate strategies to address matters related to water shortages and finally enhance sustainable adaptation to changing context.

In Vivo Reconstruction of the Acetabular Bone Defect by the Individualized Three-Dimensional Printed Porous Augment in a Swine Model.

METHODS: As an acetabular bone defect model created in Bama miniswine, an augment individually fabricated by 3D print technique with Ti6Al4V powders was implanted to repair the defect. Nine swine were divided into three groups, including the immediate biomechanics group, 12-week biomechanics group, and 12-week histological group. The inner structural parameters of the 3D printed porous augment were measured by scanning electron microscopy (SEM), including porosity, pore size, and trabecular diameter. The matching degree between the postoperative augment and the designed augment was assessed by CT scanning and 3D reconstruction. In addition, biomechanical properties, such as stiffness, compressive strength, and the elastic modulus of the 3D printed porous augment, were measured by means of a mechanical testing machine. Moreover, bone ingrowth and implant osseointegration were histomorphometrically assessed. RESULTS: In terms of the inner structural parameters of the 3D printed porous augment, the porosity was 55.48 ± 0.61%, pore size 319.23 ± 25.05 µm, and trabecular diameter 240.10 ± 23.50 µm. Biomechanically, the stiffness was 21464.60 ± 1091.69 N/mm, compressive strength 231.10 ± 11.77 MPa, and elastic modulus 5.35 ± 0.23 GPa, respectively. Furthermore, the matching extent between the postoperative augment and the designed one was up to 91.40 ± 2.83%. Besides, the maximal shear strength of the 3D printed augment was 929.46 ± 295.99 N immediately after implantation, whereas the strength was 1521.93 ± 98.38 N 12 weeks after surgery (p = 0.0302). The bone mineral apposition rate (µm per day) 12 weeks post operation was 3.77 ± 0.93 µm/d. The percentage bone volume of new bone was 22.30 ± 4.51% 12 weeks after surgery. CONCLUSION: The 3D printed porous Ti6Al4V augment designed in this study was well biocompatible with bone tissue, possessed proper biomechanical features, and was anatomically well matched with the defect bone. Therefore, the 3D printed porous Ti6Al4V augment possesses great potential as an alternative for individualized treatment of severe acetabular bone defects.

IL-6 and INF-γ levels in patients with brucellosis in severe epidemic region, Xinjiang, China.

BACKGROUND: The incidence of brucellosis, which is caused by the Brucella species of bacteria, is rapidly rising worldwide; however, few studies have investigated the immune response to this pathogen and clinical biochemical features. In this paper, we examined the levels of various cytokines and inflammatory factors as well as clinical course characteristics in patients with brucellosis, in order to provide evidence for the diagnosis, assessment, and prognosis of this infectious disease. METHODS: A total of 191 brucellosis inpatients (50 acute cases and 141 chronic cases), as well as 60 healthy control subjects, were included in the analysis. We investigated changes in the levels of six cytokines (IL-4, IL-6, IL-10, IL-17, TNF-α, INF-γ) and related clinical biochemical markers in patients with acute and chronic brucellosis in Xinjiang, China. Possible factors were statistically analyzed using the t test, χ2 test, z test and a multivariate logistic stepwise regression test. RESULTS: We found that IL-4, IL-6, IL-10, IL-17, IFN-γ, and TNF-α levels were higher in those with brucellosis than in controls (P <  0.05). With regard to disease progression, procalcitonin (PCT) and C-reactive protein (CRP) levels were significantly higher in those with an acute infection compared to chronic cases (P <  0.05). We found that the expression of all six cytokines tested was closely related to the degree of brucellosis using univariate logistic regression; however, only IL-6 and INF-γ levels were independent factors associated with the severity of brucellosis. CONCLUSIONS: Assessing cytokine levels in patients with acute and chronic brucellosis is not only useful for detecting the immune response, but can also be indicative of the severity of brucellosis. In particular, we propose IL-6 and INF-γ levels may be useful independent predictive factors in the clinical evaluation and diagnosis of brucellosis.

Optimization of microbial inactivation of shrimp by dense phase carbon dioxide.

Microbial inactivation of Litopenaeus vannamei by dense phase carbon dioxide (DPCD) treatment was investigated and neural network was used to optimize the process parameters of microbial inactivation. The results showed that DPCD treatment had a remarkable bactericidal effect on microorganism of shrimp. A 3×5×2 three-layer neural network model was established. According to the neural network model, the inactivation effect was enhanced with pressure, temperature and exposure time increasing and temperature was the most important factor affecting microbial inactivation of shrimp. Cooked appearance of shrimp by DPCD treatment was observed and seemed to be more positively acceptable by Chinese diet custom. Therefore, color change of shrimp by DPCD treatment could have a positive effect on quality attributes. Moderate temperature 55 °C with 15 MPa for 26 min treatment time achieved a 3.5-log reduction of total aerobic plate counts (TPC). The parameters combination might be appropriate for shrimp process by DPCD.

Characterization, expression and localization of S-adenosylhomocysteine hydrolase from amphioxus Branchiostoma belcheri tsingtaunese.

A cDNA clone encoding AmphiSAHH [amphioxus SAHH (S-adenosylhomocysteine hydrolase)] protein was isolated from a cDNA library from the gut of Branchiostoma belcheri tsingtaunese. It contained a 1305 bp open reading frame corresponding to a deduced protein of 434 amino acid residues, with a predicted molecular mass of approx. 47.8 kDa. Phylogenetic analysis showed that AmphiSAHH and sea-urchin SAHH joined together and positioned at the base of the vertebrate SAHH clade, suggesting that both AmphiSAHH and sea-urchin SAHH might share some characteristics of the archetype of vertebrate SAHH proteins. The genomic DNA sequence of AmphiSAHH contained eight exons and seven introns, which was similar to B. floridae and sea-urchin SAHH exon/intron organization. Sequence comparison suggested the evolutionary appearance of the ten exon/nine intron organization of SAHH genes after the split of invertebrates and vertebrates, after which it has been highly conserved. AmphiSAHH has been successfully expressed in Escherichia coli and purified. Western blotting confirmed that the enzyme has a native molecular mass of approx. 48 kDa, and the catalytic activities and NAD(+)/NADH binding affinity of recombinant AmphiSAHH were measured. Immunohistochemistry analysis showed that SAHH was strongly expressed in hepatic caecum, gill, spermary and ovary of amphioxus.